khatipo at ZAPuchicago.edu
Sat Aug 11 23:46:44 EST 2001
As far as I remember, most textbooks say that SDS denatures proteins by
"stretching" them out, which means that the whatever native conformation is
destroyed completely (I am talking about boiling in SDS). However, remember,
that residual disulfide bonds may determine the shape of the denatured
molecule. S=S bonds may not be destroyed completely by BME or DTE (the
latter being more effective), or if destroyed, may reform under oxidizing
conditions later on, unless you do not block SH with thiol reagents like
In approximation, there is one SDS molecule per 2 aa. I believe the
detergent denatures (stretches out) the protein not because of its charge,
but due to hydrophobic-hydrophilic interactions. An important feature is
that coating of the protein chain with SDS gives the proteins a similar net
negative charge that is (ideally) uniform, and constant charge-to mass
ratio. The letter allows separation of the denatured proteins during
SDS-PAGE exclusively by their size(mass) due to sieving effect of the gel,
because intrinsic mobility of proteins is (due to similar charge-to-mass
ratio) the same for most proteins. The exceptions of the latter are proteins
with extreme pI, non-uniformly charged molecules, and short peptides.
Thus, it seems likely that SDS-denatured proteins are not exactly like a
"ball of wool unrolled", but rather loose and changing in form bundle or
lump. There is certainly a chance that this thread will be completely
unwound and become a linear stretched rod, but most of the long-chain
proteins will have a irregular bundle-like shape. Shorter peptides will
stretch completely though.
Does this make sense?
"Michael Witty" <mw132 at mole.bio.cam.ac.uk> wrote in message
news:Pine.SGI.4.33.0108091040010.4361379-100000 at mole.bio.cam.ac.uk...
> Dear All,
> there is a debate in my lab about the extent to which SDS
> denatures protein. I think that SDS sticks, but the gross shape of the
> protein is not altered very much (the protein still looks like a ball of
> wool). My collegue thinks that the protein is dramatically unfolded and
> looks like a linear object, or a ball of wool unrolled. What do we think?
> Regards, Mike.
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